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Published on: May 10, 2017
[A preliminary study on the expression and biological function of recombinant human SCF-TPO fusion protein]
Nan Liu1, Yong-zhi Xi, Si-qi Guo
1Department of Immunology, Affiliated Hospital of AMMS. Laboratory of Immunoassary of NCBA, Beijing 100039, China.
Objective:
To study the expression of recombinant human SCF-TPO fusion protein and its biological function.
Methods:
Four primers were designed according to known sequences of TPO and SCF. The functional amino acid domains of TPO and SCF were amplified by RT-PCR from fetus hepatocytes, respectively. The expression plasmid pET32a/SCF-TPO was constructed by VOE gene fusion technique and expressed in E. coli BL21(DE3) plysS as inclusion body after isopropyl-beta-D-1-thiogalactopyranoside (IPTG) induction. The fusion protein was tested by SDS-PAGE and Western blot. The biological functions of SCF-TPO fusion protein in MO7e cells was investigated by MTT method after purification with metal chelating chromatography.
Results:
The high expression SCF-TPO fusion protein was obtained, reaching up to 30% of the total cellular protein. Western blot verified the correct fusion expression and MTT results showed the growth promoting effect of the SCF-TPO fusion protein on MO7e cells, with a higher promoting activity at 100 ng/ml.
Conclusions:
Expressed SCF-TPO fusion protein after renaturation has biological activity in promoting the proliferation of MO7e cells.
